During lactation, the PTHrP gene is highly expressed in mammary epithelial cells (MEC), and milk is the most abundant source of PTHrP in nature. Although the exact function of PTHrP during lactation remains unknown, it has been suggested that mammary-derived PTHrP circulates and participates in the regulation of calcium mobilization from the maternal skeleton and/or transport of calcium across MEC and into milk. In order to test this hypothesis, we used the cre/lox recombinase system to generate mice in which the PTHrP gene was disrupted only in MEC and only during lactation. Preliminary analysis of these mice suggested that withdrawal of PTHrP led to degeneration of lobuloalveolar structures in the lactating mammary gland and MEC apoptosis. This phenotype resembles the initial stages of mammary involution upon weaning, and analysis of PTHrP gene expression demonstrates that it is normally downregulated co- incident with the onset in involution. Therefore, our central hypothesis is that PTHrP serves two functions during lactation. The first is to act locally in a paracrine, autocrine or intracrine fashion to promote the survival of MEC during lactation. The second is to act systemically to participate in the regulation of maternal calcium homeostasis and to promote the mobilization of calcium from the maternal skeleton and its transport into milk. Reflecting this potential dual role, we propose two broad specific aims. Specific Aim 1 seeks to examine the possibility that PTHrP supports MEC viability during lactation. We will characterize the nature of MEC cell death in the cre-lox model, examine the possibility that PTHrP acts via a nuclear pathway and explore the potential relationships between PTHrP and the RANKL/RANK/OPG system. Specific Aim 2 seeks to determine if PTHrP regulates calcium transport and/or mobilization during lactation. We will use a cell culture system to investigate the potentially interrelated effects of PTHrP and the calcium-sensing receptor on calcium transport across MEC and will use the cre-lox model to determine if mammary-derived PTHrP, potentially interacting with IL-6, regulates maternal calcium and/or bone metabolism during lactation. We think the experiments outlined in this proposal should offer insight into the role of PTHrP during lactation.